The integration and expression of murine leukemia virus DNA in mouse cells will be studied using recombinant DNA cloning techniques. Permanent mouse fibroblast lines clonally infected with Moloney murine leukemia virus (M-MuLV) have been prepared, and the M-MuLV-specific integrations will be identified in each cell line by hybridization to Southern gel blots of infected cell DNA cleaved with Eco RI, an enzyme which does not cleave M-MuLV DNA. M-MuLV DNA copies which are active and inactive in transcription will be identified by DNase I digestion of nuclei. Eco RI fragments containing the integrated M-MuLV DNA copies will be cloned in bacteriophage lambda cloning vectors, and both the viral sequences and the adjacent cellular sequences will be studied. Analyses will include determination of viral sequence organization, hybridization to uninfected cell DNA, tests for homology among the clones, primary sequence analysis and biological assays by DNA transfection. These experiments may indicate if any specificity for the integration sites in the cell DNA exists. Regulatory elements affecting the expression of the viral DNA may also be identified. Recombinant DNA clones containing endogenous M-MuLV-related retroviral information will also be studied in a similar manner. These studies may yield information on the events leading to the integration of endogenous virus sequences in the germ line, and will allow us to test if integration of these endogenous viruses was similar to that for exogenously infecting M-MuLV in fibroblasts.